Let-7d inhibits proliferation, migration and invasion of osteosarcoma cells by targeting rhotekin gene / 第二军医大学学报

ABSTRACT

Objective To investigate the expression level of let-7d in osteosarcoma tissues, and to explore the effects of let-7d and its target gene on proliferation, migration and invasion of human osteosarcoma cell line U2OS cells. Methods Tumor and adjacent tumor tissues of 25 patients with osteosarcoma undergoing exairesis in our department were collected from 2010 to 2015. The distance from adjacent tumor tissues to tumor margin was greater than 5 cm. The expressions of let-7d were detected by qPCR in the tumor and adjacent tumor tissues. A U2OS cell model stably overexpressing let-7d was constructed, and the expression of let-7d was confirmed by qPCR. The U2OS cells transfected with pCDH empty virus vector were used as control cells. CCK-8 assay, scratch assay and Transwell assay were used to determine the proliferation, migration and invasion of the cell after transfection, respectively. A downstream target gene of let-7d was identified based on microRNA target gene prediction software and luciferase activity assay, and the expression of the target gene was detected in U2OS cells overexpressing let-7d. Effects of the target gene on the cell proliferation, migration and invasion were analyzed by small interfering RNA technology. Results The expression level of let-7d was significantly lower in the osteosarcoma tissues than that in the adjacent tumor tissues (P<0.01). The expression level of let-7d was significantly lower in U2OS cells compared with human osteoblast cell line hFOB1.19 cells (P<0.01). Compared with the control group, overexpression of let-7d significantly reduced the proliferation, migration and invasion of U2OS cells (all P<0.05). MicroRNA target gene prediction software and luciferase activity assay showed that Rhotekin (RTKN) gene was a direct target gene of let-7d. Compared with the control group, overexpression of let-7d significantly reduced mRNA expression level of RTKN in the U2OS cells (P<0.01). Down-regulation of RTKN significantly inhibited the proliferation, migration and invasion of U2OS cells (all P<0.05). Conclusion Let-7d inhibits proliferation, migration and invasion of osteosarcoma cells by targeting RTKN, which indicates let-7d may be a novel candidate biological therapeutic target for osteosarcoma.